This invention relates to detection of charged and neutral molecules and fragments, and more particularly to the detection of such species generated in a mass spectrometer.
Mass spectrometry is an analytical technique for the determination of molecular weights, the identification of chemical structures, the determination of the composition of mixtures, and qualitative elemental analysis. In operation, a mass spectrometer generates ions of sample molecules under investigation (the analyte), separates the ions according to their mass-to-charge ratio, and measures the relative abundance of each ion.
Mass spectrometry involves introducing a sample presentation apparatus into the mass spectrometer, volatilizing and ionizing the analyte, accelerating the ionized analyte toward a detector by exposing the ions to an electric and/or magnetic field, and analyzing the data to determine the mass to charge ratio (m/q) of specific analyte ions. If the analyte remains intact throughout this process, data obtained will correspond to a molecular weight for the entire intact analyte ion. Typically, however, and especially for the case of larger biological analytes, it is beneficial to obtain data corresponding to the molecular weight of various fragments of the analyte. It is also desirable to obtain data which only corresponds to the pure analyte, even when impurities are present.
Mass spectrometry techniques include quadrupole, magnetic sector and time of flight (TOF) methods. Time of flight mass spectrometry (TOF) is a technique that separates different ion mass by a time coordinate. First, ions are accelerated across a given voltage so that they will attain common kinetic energies. Thus, ions of different mass/charge ratio (m/q) will attain different velocities. If these ions are allowed to drift, they will spread out in space, and the lightest (and fastest) ions will arrive at the detector first. A time-sensitive detection system can be used to reconstruct a mass spectrum. TOF mass spectrometers are advantageous because they are relatively simple, inexpensive instruments with virtually unlimited mass-to-charge ratio range. TOF mass spectrometers have potentially higher sensitivity than scanning instruments because they can record all the ions generated from each ionization event. TOF mass spectrometers are particularly useful for measuring the mass-to-charge ratio of large organic molecules where conventional magnetic field mass spectrometers can lack sensitivity. TOF mass spectrometers are shown, for example, in U.S. Pat. Nos. 5,045,694 and 5,160,840.
TOF mass spectrometers include an ionization source for generating analyte ions. The ionization source contains one or more electrodes or electrostatic lenses for accelerating and properly directing the ion beam. In the simplest case the electrodes are grids. A detector is positioned a predetermined distance from the final grid for detecting ions as a function of time.
Matrix-assisted laser desorption/ionization (MALDI) is a technique to volatilize and ionize biological molecules in a mass spectrometer which uses TOF techniques. MALDI involves surrounding a biomolecule with a matrix material. A laser beam, tuned to a frequency where the matrix material absorbs, is targeted on the matrix material. The laser transfers sufficient energy to volatilize a small portion of the matrix material. A small number of analyte molecules are thus carried along with the matrix material into the vapor phase in the mass spectrometer.
TOF systems have used electron multiplier detectors of several types, including box-and-grid, Venetian blind, magnetic strip and single channel electron multipliers. In these, rise times can be too long to resolve close-lying mass peaks. Electronic gating has been used to measure only a single m/q for each ion group, with the interval for detection advanced stepwise for subsequent ion-generating pulses. Covering the entire mass range for a given sample requires a very large number of pulses and consequently long measurement times. As mass spectrometry is a destructive analytical technique, precious samples were sacrificed for these measurements as well.
A microchannel plate (MCP) detector is a wafer-like lead glass microchannel device with superior timing resolution ( less than 1ns rise time) that can alleviate these problems. MCP allows the detection of all m/q values in a single ion-generating pulse. This advance motivated parallel advances in other areas of TOF, such as mass resolution. Further background on MCP detectors is found in J. L. Wiza, Nucl. Instr. Meth., 162, (1979) p 587.
TOF is used in biomedical research and clinical applications, commonly through laser desorption ionization (MALDI) and electrospray ionization techniques. However, very low detector efficiency for such ions can become a limiting factor. Ion detectors not suited for efficient detection of high mass biomolecular ions hinder accurate mass analysis of proteins and other biomolecules of importance to biochemistry, modem biology, and medical science.
Although the timing problem has been improved with the MCP, other detector limitations can limit TOF mass spectrometry. These drawbacks include poor sensitivity to high-mass ions and inadequate dynamic range. One way to increase detector sensitivity is to increase the incident ion postacceleration voltage to 25 or 35 kV or more to increase ion yields and detector signal levels.
The invention provides detection of molecules in mass spectroscopy instruments. In embodiments, significant improvements in detection sensitivity can result from the use of a repeller grid placed in the vicinity of the detector face, particularly in combination with a coating of low work function material on the detector face, the coating being of a relatively low density. A further improvement in signal can be produced by utilizing a negative interplate bias when detectors are used in a tandem configuration. The detection signal generated by detectors utilizing this combination of features is superior to that of detectors without either feature, or with each feature utilized independently. The combination of features provides a synergistic effect in producing a detection signal.
The detection system offers superior detection sensitivity for high mass ions over conventional detection systems. The detection system obviates complex detector manufacturing methods considered necessary for obtaining acceptable sensitivity. Moreover, the detection system, when used in a tandem configuration, further improves detection efficiency of high mass ions by reducing the interfering light mass matrix ion signal which is found in MALDI techniques.
Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. Although methods and materials similar or equivalent to those described herein can be used in the practice or testing of the present invention, suitable methods and materials are described below. All publications, patent applications, patents, and other references mentioned herein are incorporated by reference in their entirety. In case of conflict, the present specification, including definitions, will control. In addition, the materials, methods, and examples are illustrative only and not intended to be limiting.
Other features and advantages of the invention will be apparent from the following detailed description, and from the claims.